DESCRIPTION (adapted from application abstract): This is a study of host- pathogen interactions at the molecular level, specifically the structural functional relationship of bacterial attachment to host epithelial cells. Urinary tract infections (UTIs) affect a large proportion of the world population and account for significant morbidity and high medical costs. Escherichia coli is the etiologic agent in the majority of all UTIs, including cyctitis. Other members of the Enterobacteriaceae family, including Klebsiella species and Proteus mitabilis, and Gram-positive organisms, including Staphylococcus saprophyticus and opportunistic enterococci species, are also associated with UTIs. To act as pathogens, these various bacteria must first adhere to host mucosal surfaces within the urinary tract. Without the ability to specifically adhere to host tissues, these pathogens would be readily expelled from the host and disease would not occur. Bacterial attachment can initiate a cascade of molecular crosstalk between bacterial and host cells that can directly influence the course of an infection. Virtually all uropathogenic strains of E. coli and most members of the Enterobacteriaceae family encode filamentous surface adhesive organelles called type 1 pili. These structures can mediate bacterial attachment to bladder epithelial cells by interacting with host receptor complexes comprised of integral membrane proteins known as uroplakins. The candidate proposes to use type 1-piliated uropathogenic E. coli as a model to study the structural basis of bacterial attachment and the functional consequences of the attachment event. The specifics of the interactions between type 1 pili and uroplakin receptor complexes will be examined. In addition, the bacterial and host factors that come into play as a consequence of bacterial adherence and influence the pathogenesis of UTIs will be analyzed at both the cellular and molecular level. This work will lead to enhanced understanding of pathogenic processes both within the urinary tract and at other sites of microbial entry and will facilitate the development of novel antimicrobial therapeutics and vaccines. The specific aims are to: 1.Characterize interactions between type 1 pili and the uroplakin receptor complexes that coat the lumenal surface of the bladder. Mutational analysis of recombinant uroplakin protein complexes and of type 1 pili along with biochemical, microscopic, and crystallographic techniques will provide detailed insight into the host-pathogen interactions involved in the establishment of UTIs by uropathogenic E. coli. 2.Investigate virulence factors and mechanisms that enable uropathogenic E. coli to invade, disseminate, and persist within the bladder following the initial attachment event mediated by type 1 pili. The possibility that recurrent UTIs may, in some cases, be a manifestation of a lingering chronic infection will be tested. 3.Elucidate host responses to uropathogenic E. coli, including the identification and characterization of innate and adaptive host defenses that function to limit and clear invading bacteria from the urinary tract. Cell culture systems and a murine cystitis model will be used to delineate vital constituents of an effective host response to infection by type 1-piliated E. coli.